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A novel disulfide bond-mediated cleavable RGD-modified PAMAM nanocomplex containing nuclear localization signal HMGB1 for enhancing gene transfection efficiency.

ABSTRACT: Background:Polyamidoamine (PAMAM) dendrimers modified by polyethylene glycol (PEG) have frequently been investigated as a delivery carrier for gene therapy. However, modification of PAMAM with PEG using covalent linkage significantly reduces the cellular uptake rate and the transfection efficiency. How to conquer these barriers becomes a burning question in gene delivery. Materials and methods:The present study constructed an effective disulfide bond-mediated cleavable RGD modified gene delivery system to overcome the aforementioned limitations. The disulfide bond was introduced between PAMAM dendrimers and PEG chains to realize the cleavage of PEG from the carrier system, whereas the arginine-glycine-aspartate (RGD) peptide was expected to promote the cellular uptake rate. A high mobility group Box 1 (HMGB1) protein containing nuclear localization signal (NLS) was simultaneously introduced to further promote gene expression efficiency. A pDNA/HMGB1/PAMAM-SS-PEG-RGD (DHP) nanocomplex was prepared via electrostatic interaction and characterized. Results:The results showed that DHP generated small particles and was able to condense and protect pDNA against degradation. In addition, the RGD peptide could significantly promote the cellular uptake of a nanocomplex. Intracellular trafficking and in vitro expression study indicated that the DHP nanocomplex escaped from lysosomes and the disulfide bonds between PAMAM and PEG cleaved due to the high concentration of GSH in the cytoplasm, pDNA consequently became exclusively located in the nucleus under the guidance of HMGB1, thereby promoting the red fluorescence protein (RFP) expression. Importantly, an in vivo antitumor activity study demonstrated that the DHP nanocomplex had higher antitumor activity than any other reference preparation. Conclusion:All these results confirm that DHP could be a new strategy for improving the transfection and expression efficiency in gene delivery.

SUBMITTER: Li J

PROVIDER: S-EPMC6228086 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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A novel disulfide bond-mediated cleavable RGD-modified PAMAM nanocomplex containing nuclear localization signal HMGB1 for enhancing gene transfection efficiency.

Li Ji J Han Yuting Y Lu Yue Y Song Baohui B Zhao Ming M Hu Haiyang H Chen Dawei D

International journal of nanomedicine 20181106

<h4>Background</h4>Polyamidoamine (PAMAM) dendrimers modified by polyethylene glycol (PEG) have frequently been investigated as a delivery carrier for gene therapy. However, modification of PAMAM with PEG using covalent linkage significantly reduces the cellular uptake rate and the transfection efficiency. How to conquer these barriers becomes a burning question in gene delivery.<h4>Materials and methods</h4>The present study constructed an effective disulfide bond-mediated cleavable RGD modifie ...[more]

PMID: 30464464

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Project description:The protein secretory pathway must maintain homoeostasis while producing a wide assortment of proteins in different conditions. It is also used extensively to produce many useful proteins in biotechnology. As such, secretory pathway dysfunction can be highly detrimental to the cell, resulting in the molecular basis for many human diseases, and can drastically inhibit product titers in biochemical production. Because the secretory pathway is a highly-integrated, multi-organelle system, dysfunction can happen at many levels and dissecting the root cause can be challenging. To better understand some of these dysfunctions, we measured multiple systems-level states of the cell (physiology, transcriptome, metabolism) while secreting a small protein (insulin precursor) or a large protein (?-amylase). This was carried out in the presence and absence of HAC1, a key transcription factor in maintaining secretory homeostasis. Clear trends in cellular stress were apparent across multiple data resulting from our perturbations. In particular, processes involving (1) degradation of protein / recycling amino acids, (2) overall transcription/translation repression, and (3) oxidative stress. Apparent runaway oxidative radical production was explained by a thermodynamic model that we put forward for disulfide formation in the endoplasmic reticulum. This model predicts that balancing the relative rates of protein folding and disulfide bond formation are key to easing oxidative stress. These predictions have direct implications in how to engineer a broad range of recombinant proteins for secretion and provide potential hypotheses for the root causes of several secretory-associated diseases. Yeast strains were constructed that produce and secrete (a) IP or (b) ?-amylase and were compared to yeast strains containing (c) an empty vector in both wild-type and HAC1 deletion backgrounds. These strains are named WN (WT with empty vector), WI (WT secreting IP), WA (WT secreting ?-amylase), dN (?hac1 with empty vector), dI (?hac1 secreting IP), and dA (?hac1 secreting ?-amylase). Strains were characterized in batch fermentation and samples were taken in mid-exponential phase. Triplicate fermentations were carried out for each strain.

Dataset Information

A novel disulfide bond-mediated cleavable RGD-modified PAMAM nanocomplex containing nuclear localization signal HMGB1 for enhancing gene transfection efficiency.

Publications

A novel disulfide bond-mediated cleavable RGD-modified PAMAM nanocomplex containing nuclear localization signal HMGB1 for enhancing gene transfection efficiency.

Similar Datasets

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